Oil seed meal preparation

ABSTRACT

Canola oil seeds are treated for the production of a canola oil seed meal for recovery of canola protein isolates therefrom. The canola oil seeds are heat-treated to inactivate myrosinases and other enzymes and dehulled prior to crushing dehulled canola oil seeds and removing oil therefrom and to provide the canola oil seed meal.

FIELD OF INVENTION

The invention is directed to the preparation of oil seed meal for therecovery of protein therefrom.

BACKGROUND OF THE INVENTION

In copending U.S. patent application Ser. Nos. 10/137,391 filed May 3,2002 (WO 02/089597) and 10/476,230 filed Jun. 9, 2004, all assigned tothe assignee hereof and the disclosures of which are incorporated hereinby reference, there is described a process for producing a proteinisolate of high purity, containing at least about 100 wt % protein whendetermined by the Kjeldahl or equivalent method as percent nitrogen (N)and multiplied by a conversion factor of 6.25. As used herein, the term“protein content” refers to the quantity of protein in the proteinisolate expressed on a dry weight basis. In the aforementioned U.S.Patent Applications, the protein isolate is made by a process in whichoil seed meal is extracted with a food grade salt solution, theresulting protein solution, after an initial treatment with a colourantadsorbent, if desired, is concentrated to a protein content of at leastabout 200 g/L, and the concentrated protein solution is diluted inchilled water to form protein micelles, which are allowed to settle toform an aggregated, coalesced, dense amorphous, sticky gluten-likeprotein isolate mass, termed “protein micellar mass” or PMM, which isseparated from residual aqueous phase and may be used as such or dried.

In one embodiment of the process described above and as specificallydescribed in U.S. patent application Ser. Nos. 10/137,391 and10/476,230, the supernatant from the PMM settling step is processed torecover a protein isolate comprising dried protein from wet PMM andsupernatant. This procedure may be effected by initially concentratingthe supernatant using ultrafiltration membranes, mixing the concentratedsupernatant with the wet PMM and drying the mixture. The resultingcanola protein isolate has a high purity of at least about 90 wt %,preferably at least about 100 wt %, protein (N×6.25).

In another embodiment of the process described above and as specificallydescribed in application Ser. Nos. 10/137,391 and 10/476,230, thesupernatant from the PMM settling step is processed to recover a proteinfrom the supernatant. This procedure may be effected by initiallyconcentrating the supernatant using ultrafiltration membranes and dryingthe concentrate. The resulting canola protein isolate has a high purityof at least about 90 wt %, preferably at least about 100 wt %, protein(N×6.25).

The procedures described in the aforementioned US Patent Applicationsare essentially batch procedures. In copending U.S. patent applicationSer. No. 10/298,678 filed Nov. 19, 2002 (WO 03/043439), assigned to theassignee hereof and the disclosures of which are incorporated herein byreference, there is described a continuous process for making canolaprotein isolates. In accordance therewith, canola oil seed meal iscontinuously mixed with a salt solution, the mixture is conveyed througha pipe while extracting protein from the canola oil seed meal to form anaqueous protein solution, the aqueous protein solution is continuouslyseparated from residual canola oil seed meal, the aqueous proteinsolution is continuously conveyed through a selective membrane operationto increase the protein content of the aqueous protein solution to atleast about 200 g/L while maintaining the ionic strength substantiallyconstant, the resulting concentrated protein solution is continuouslymixed with chilled water to cause the formation of protein micelles, andthe protein micelles are continuously permitted to settle while thesupernatant is continuously overflowed until the desired amount of PMMhas accumulated in the settling vessel. The PMM is removed from thesettling vessel and may be dried. The PMM has a protein content of atleast about 90 wt % (N×6.25), preferably at least about 100 wt %.

The meal which is extracted at the initial step in the preparation ofthe protein isolate contains a number of components which can contributeto the taste and colour of the protein isolate. For example, there arehull particles that contain certain phenolic compounds which may leachinto the extract. Such phenolic compounds are prone to oxidation to formcoloured compounds.

Other components which may contribute to the quality of the meal and itsproducts are glucosinolates and the products of their degradation.Degradation of glucosinolates is catalyzed by degrative enzymes calledmyrosinases, which break down glucosinolates into isothyocyanates,thiocyanates, nitriles and elemental sulfur. The degradation products ofglucosinolates reduce the value of glucosinolate containing plants whenused as food for humans or for feeding animals.

Canola is also known as rapeseed or oil seed rape.

SUMMARY OF THE INVENTION

In the present invention, canola oil seeds, are subjected to heattreatment to inactivate the myrosinases and to dehulling prior tocrushing the dehulled oil seeds to remove oil therefrom. The procedureminimizes the presence of components in the meal which adversely affectcolour and taste of the protein isolate derived from the oil seed mealusing the processes described above. The heat treatment procedureprovided herein also may be used to deactivate other enzymes which maybe present in the oil seed.

The inactivation of myrosinases and other enzymes present in the canolaoil seeds may be effected in any convenient manner consistent withinactivation of the enzymes. Most conveniently, the inactivation iscarried out using steam at approximately 90° C. for a minimum of 10minutes, although other temperatures, times and procedures may be used,for example, the use of infra-red, microwave or radio frequencytreatment. The important feature is that the enzymes, including themyrosinases, are inactivated.

Accordingly, in one aspect of the present invention, there is provided amethod of forming a canola oil seed meal, which comprises heat treatingthe canola oil seeds to deactivate enzymes therein, dehulling the canolaoil seeds, and removing canola oil from the heat treated and dehulledoil seeds to provide the canola oil seed meal.

The canola oil seed meal produced by the process then may be processedto recover canola protein isolates therefrom having a protein content ofat least about 90% by weight (N×6.25), preferably at least 100% byweight. The canola protein isolation procedure used preferably is one ofthose described in the aforementioned US Patent Applications.

BRIEF DESCRIPTION OF DRAWINGS

FIG. 1 is a process flow chart of a preparative procedure for obtaininga dehulled and defatted canola oil seed in accordance with one preferredembodiment of the invention;

FIG. 2 is a process flow chart of a preparative procedure for obtaininga dehulled and defatted canola oil seed in accordance with a lesspreferred embodiment of the invention;

FIG. 3 is a flow chart for the preparation of a canola protein isolatefrom the dehulled and defatted canola oil seed prepared according to theprocedure of FIG. 1 or FIG. 2; and

FIG. 4 is a graphical representative of temperature profiles for heattreatment of canola oil seed and dehulled meat fractions.

GENERAL DESCRIPTION OF INVENTION

The present invention is concerned with the processing of canola oilseeds to produce a canola oil seed meal from which canola proteinisolates can be prepared.

The process involves heat treatment of the canola oil seeds toinactivate myrosinase enzymes and other enzymes present in the seeds anddehulling of the seeds. The dehulling may be effected following heattreatment or before heat treatment. The processed seeds then aresubjected to an oil removal step to leave a canola oil seed meal.

The heat treatment may conveniently be effected at approximately 90° C.for a minimum of about 5 minutes, preferably about 10 minutes, usingsteam heating. As mentioned above, other temperatures, times andprocedures may be used, such as infra-red, microwave or radio frequencytreatment. Following heat treatment, the oil seeds are usually cooled toambient temperature for further processing.

In one embodiment of the invention, illustrated in FIG. 1, canola oilseeds are first inactivated in a cooker at about 90° C. for about 10minutes by steam injection. The inactivated oil seeds then are cooled toambient temperature, such as by employing a fluid bed dryer. The cooleddeactivated canola oil seeds then are forwarded to a cracking millwherein canola hulls are cracked and the cracked hulls are separatedfrom canola meats, such as by air aspiration. The canola meats areseparated into a larger (overs) fraction and a smaller (unders)fraction, such as by the use of a vibratory screen. In the illustratedexample of FIG. 1, a 14 mesh screen is used for the separation step.

The overs fraction tends to have more residual uncracked hullsassociated therewith and generally is recycled to the cracking millseveral times to remove residual hulls. Once the overs fraction has beendehulled, it can be processed for the recovery of canola oil andproduction of canola oil seed meal, by flaking the meat and effectingsolvent extraction of the flakes. The recovered meal usually isdesolventized.

The unders fraction is processed to remove residual hulls, such as byair aspiration. Once the unders fraction has been dehulled, it can beprocessed for the recovery of canola oil and production of canola oilseed meal, by flaking the meat and effecting solvent extraction of theflakes. The remaining meal usually is desolventized. The canola meatovers and unders may concurrently be combined prior to the flaking step.

In another embodiment of the invention illustrated in FIG. 2, enzymeinactivation is effected after dehulling. The canola oil seed meal isfed to a cracking mill wherein canola hulls are cracked and the crackedhulls are separated from canola meats. The canola meats are separatedinto a larger (overs) fraction and a smaller (unders) fraction, such asby the use of a vibratory screen. The overs fraction tends to have moreresidual uncracked hulls associated therewith and generally is recycledto the cracking mill several times to remove residual hulls.

Each of the overs and unders fractions is subjected to inactivation in acooker at about 90° C. for 10 minutes by steam injection. Theinactivated fractions then are separately cooled, such as by utilizing afluid bed dryer.

The cooled fractions then are processed for the recovery of canola oiland production of canola oil seed meal. The overs fraction is subjectedto flaking, residual hull removal, such as by air aspiration, andsolvent extraction of the flakes. The remaining meal may bedesolventized.

The unders fraction is subjected to flaking and the flakes are subjectedto solvent extraction. The residual meal may be desolventized.

The residual meal prepared by these procedures is further processed torecover canola protein isolate therefrom using the procedure describedin the aforementioned US patent applications, as described in furtherdetail below.

The respective PMM-derived canola protein isolate andsupernatant-derived canola protein isolate may be isolated from canolaoil seed meal by either a batch process or a continuous process or asemi-continuous process as generally described in the aforementionedUnited States patent applications.

The initial step of the process of providing the canola protein isolatesinvolves solubilizing proteinaceous material from canola oil seed meal.The proteinaceous material recovered from canola seed meal may be theprotein naturally occurring in canola seed or the proteinaceous materialmay be a protein modified by genetic manipulation but possessingcharacteristic hydrophobic and polar properties of the natural protein.The canola meal may be any canola meal resulting from the removal ofcanola oil from canola oil seed with varying levels of non-denaturedprotein, resulting, for example, from hot hexane extraction or cold oilextrusion methods. The removal of canola oil from canola oil seedusually is effected as a separate operation from the protein isolaterecovery procedure described herein.

Protein solubilization is effected in accordance with the presentinvention by using a salt solution. The salt usually is sodium chloride,although other suitable salts, such as potassium chloride and calciumchloride, may be used. The salt solution has an ionic strength of atleast about 0.10, preferably at least about 0.15, to enablesolubilization of significant quantities of protein to be effected. Asthe ionic strength of the salt solution increases, the degree ofsolubilization of protein in the oil seed meal initially increases untila maximum value is achieved. Any subsequent increase in ionic strengthdoes not increase the total protein solubilized. The ionic strength ofthe salt solution which causes maximum protein solubilization variesdepending on the oil seed meal chosen.

In view of the greater degree of dilution required for proteinprecipitation with increasing ionic strengths, it is usually preferredto utilize an ionic strength value less than about 0.8, and morepreferably a value of about 0.15 to about 0.6.

In a batch process, the salt solubilization of the protein is effectedat a temperature of at least about 5° C. and preferably up to about 35°C., preferably accompanied by agitation to decrease the solubilizationtime, which is usually about 10 to about 60 minutes. It is preferred toeffect the solubilization to extract substantially as much protein fromthe oil seed meal as is practicable, so as to provide an overall highproduct yield.

The lower temperature limit of about 5° C. is chosen sincesolubilization is impractically slow below this temperature while theupper preferred temperature limit of about 35° C. is chosen since theprocess becomes uneconomic at higher temperature levels in a batch mode.

In a continuous process, the extraction of the protein from the canolaoil seed meal is carried out in any manner consistent with effecting acontinuous extraction of protein from the canola oil seed meal. In oneembodiment, the canola oil seed meal is continuously mixed with a saltsolution and the mixture is conveyed through a pipe or conduit having alength and at a flow rate for a residence time sufficient to effect thedesired extraction in accordance with the parameters described herein.In such continuous procedure, the salt solubilization step is effectedrapidly, in a time of up to about 10 minutes, preferably to effectsolubilization to extract substantially as much protein from the canolaoil seed meal as is practicable. The solubilization in the continuousprocedure preferably is effect at elevated temperatures, preferablyabove about 35° C., generally up to about 65° C.

The aqueous salt solution and the canola oil seed meal have a natural pHof about 5 to about 6.8 to enable a protein isolate to be formed by themicellar route, as described in more detail below.

At and close to the limits of the pH range, protein isolate formationoccurs only partly through the micelle route and in lower yields thanattainable elsewhere in the pH range. For these reasons, pH values ofabout 5.3 to about 6.2 are preferred.

The pH of the salt solution may be adjusted to any desired value withinthe range of about 5 to about 6.8 for use in the extraction step by theuse of any convenient acid, usually hydrochloric acid, or alkali,usually sodium hydroxide, as required.

The concentration of oil seed meal in the salt solution during thesolubilization step may vary widely. Typical concentration values areabout 5 to about 15% w/v.

The protein extraction step with the aqueous salt solution has theadditional effect of solubilizing fats which may be present in thecanola meal, which then results in the fats being present in the aqueousphase.

The protein solution resulting from the extraction step generally has aprotein concentration of about 5 to about 40 g/L, preferably about 10 toabout 30 g/L.

The aqueous phase resulting from the extraction step then may beseparated from the residual canola meal, in any convenient manner, suchas by employing a decanter centrifuge, followed by disc centrifugationand/or filtration to remove residual meal. The separated residual mealmay be dried for disposal.

The colour of the final canola protein isolate can be improved in termsof light colour and less intense yellow by the mixing of powderedactivated carbon or other pigment adsorbing agent with the separatedaqueous protein solution and subsequently removing the adsorbent,conveniently by filtration, to provide a protein solution. Diafiltrationalso may be used for pigment removal.

Such pigment removal step may be carried out under any convenientconditions, generally at the ambient temperature of the separatedaqueous protein solution, employing any suitable pigment adsorbingagent. For powdered activated carbon, an amount of about 0.025% to about5% w/v, preferably about 0.05% to about 2% w/v, is employed.

Where the canola seed meal contains significant quantities of fat, asdescribed in U.S. Pat. Nos. 5,844,086 and 6,005,076, assigned to theassignee hereof and the disclosures of which are incorporated herein byreference, then the defatting steps described therein may be effected onthe separated aqueous protein solution and on the concentrated aqueousprotein solution discussed below. When the colour improvement step iscarried out, such step may be effected after the first defatting step.

An alternative procedure is to extract the oil seed meal with the saltsolution at a relatively high pH value above about 6.8, generally up toabout 9.9. The pH of the sodium chloride solution, may be adjusted in pHto the desired alkaline value by the use of any convenient food-gradealkali, such as aqueous sodium hydroxide solution. Alternatively, theoil seed meal may be extracted with the sodium solution at a relativelylow pH below about pH 5, generally down to about pH 3. Where suchalternative is employed, the aqueous phase resulting from the oil seedmeal extraction step then is separated from the residual canola meal, inany convenient manner, such as by employing decanter centrifugation,followed by disc centrifugation to remove residual meal. The separatedresidual meal may be dried for disposal.

The aqueous protein solution resulting from the high or low pHextraction step then is pH adjusted to the range of about 5 to about6.8, preferably about 5.3 to about 6.2, as discussed above, prior tofurther processing as discussed below. Such pH adjustment may beeffected using any convenient acid, such as hydrochloric acid, oralkali, such as sodium hydroxide, as appropriate.

The aqueous protein solution then is concentrated to increase theprotein concentration thereof while maintaining the ionic strengththereof substantially constant. Such concentration generally is effectedto provide a concentrated protein solution having a proteinconcentration of at least about 50 g/L, preferably at least about 200g/L, more preferably at least about 250 g/L.

The concentration step may be effected in any convenient mannerconsistent with batch or continuous operation, such as by employing anyconvenient selective membrane technique, such as ultrafiltration ordiafiltration, using membranes, such as hollow-fibre membranes orspiral-wound membranes, with a suitable molecular weight cut-off, suchas about 3,000 to about 100,000 daltons, preferably about 5,000 to about10,000 daltons, having regard to differing membrane materials andconfigurations, and, for continuous operation, dimensioned to permit thedesired degree of concentration as the aqueous protein solution passesthrough the membranes.

The concentrated protein solution then may be subjected to adiafiltration step using an aqueous sodium chloride solution of the samemolarity and pH as the extraction solution. Such diafiltration may beeffected using from about 2 to about 20 volumes of diafiltrationsolution, preferably about 5 to about 10 volumes of diafiltrationsolution. In the diafiltration operation, further quantities ofcontamination are removed from the aqueous protein solution by passagethrough the membrane with the permeate. The diafiltration operation maybe effected until no significant further quantities of phenolics andvisible colour are present in the permeate. Such diafiltration may beeffected using a membrane having a molecular weight cut-off in the rangeof about 3,000 to about 100,000 daltons, preferably about 5,000 to about10,000 daltons, having regard to different membrane materials andconfiguration.

An antioxidant may be present in the diafiltration medium during atleast part of the diaflitration step. The antioxidant may be anyconvenient antioxidant, such as sodium sulfite or ascorbic acid. Thequantity of antioxidant employed in the diafiltration medium depends onthe materials employed and may vary from about 0.01 to about 1 wt %,preferably about 0.05 wt %. The antioxidant serves to inhibit oxidationof phenolics present in the concentrated canola protein isolatesolution.

The concentration step and the diafiltration step may be effected at anyconvenient temperature, generally about 20° to about 60° C., preferablyabout 20 to about 30° C., and for the period of time to effect thedesired degree of concentration. The temperature and other conditionsused to some degree depend upon the membrane equipment used to effectthe concentration and the desired protein concentration of the solution.

The concentrating of the protein solution to the preferred concentrationabove about 200 g/L in this step not only increases the process yield tolevels above about 40% in terms of the proportion of extracted proteinwhich is recovered as dried protein isolate, preferably above about 80%,but also decreases the salt concentration of the final protein isolateafter drying. The ability to control the salt concentration of theisolate is important in applications of the isolate where variations insalt concentrations affect the functional and sensory properties in aspecific food application.

As is well known, ultrafiltration and similar selective membranetechniques permit low molecular weight species to pass therethroughwhile preventing higher molecular weight species from so doing. The lowmolecular weight species include not only the ionic species of the foodgrade salt but also low molecular weight materials extracted from thesource material, such as, carbohydrates, pigments and anti-nutritionalfactors, as well as any low molecular weight forms of the protein. Themolecular weight cut-off of the membrane is usually chosen to ensureretention of a significant proportion of the protein in the solution,while permitting contaminants to pass through having regard to thedifferent membrane materials and configurations.

The concentrated and optionally diafiltered protein solution may besubject to a further defatting operation, if required, as described inU.S. Pat. Nos. 5,844,086 and 6,005,076.

The concentrated and optionally diafiltered protein solution may besubject to a colour removal operation as an alternative to the colourremoval operation described above. Powdered activated carbon may be usedherein as well as granulated activated carbon (GAC). Another materialwhich may be used as a colour adsorbing agent is polyvinyl pyrrolidone.

The colour absorbing agent treatment step may be carried out under anyconvenient conditions, generally at the ambient temperature of thecanola protein solution. For powdered activated carbon, an amount ofabout 0.025% to about 5% w/v, preferably about 0.05% to about 2% w/v,may be used. Where polyvinylpyrrolidone is used as the colour adsorbingagent, an amount of about 0.5% to about 5% w/v, preferably about 2% toabout 3% w/v, may be used. The colour adsorbing agent may be removedfrom the canola protein solution by any convenient means, such as byfiltration.

The concentrated and optionally diafiltered protein solution resultingfrom the optional colour removal step may be subjected to pasteurizationto kill any bacteria which may have been present in the original meal asa result of storage or otherwise and extracted from the meal into thecanola protein isolate solution in the extraction step. Suchpasteurization may be effected under any desired pasteurizationconditions. Generally, the concentrated and optionally diafilteredprotein solution is heated to a temperature of about 55° to about 70°C., preferably about 60° to about 65° C., for about 10 to about 15minutes, preferably about 10 minutes. The pasteurized concentratedprotein solution then may be cooled for further processing as describedbelow, preferably to a temperature of about 25° to about 40° C.

Depending on the temperature employed in the concentration step, theconcentrated protein solution may be warmed to a temperature of at leastabout 20°, and up to about 60° C., preferably about 25° to about 40° C.,to decrease the viscosity of the concentrated protein solution tofacilitate performance of the subsequent dilution step and micelleformation. The concentrated protein solution should not be heated beyonda temperature above which the temperature of the concentrated proteinsolution does not permit micelle formation on dilution by chilled water.The concentrated protein solution may be subject to a further defattingoperation, if required, as described in the aforementioned U.S. Pat.Nos. 5,844,086 and 6,005,076.

The concentrated protein solution resulting from the concentration step,and optional diafiltration step, optional colour removal step, optionalpasteurization step and optional defatting step, then is diluted toeffect micelle formation by mixing the concentrated protein solutionwith chilled water having the volume required to achieve the degree ofdilution desired. Depending on the proportion of canola protein desiredto be obtained by the micelle route and the proportion from thesupernatant, the degree of dilution of the concentrated protein solutionmay be varied. With higher dilution levels, in general, a greaterproportion of the canola protein remains in the aqueous phase.

When it is desired to provide the greatest proportion of the protein bythe micelle route, the concentrated protein solution is diluted by about15 fold or less, preferably about 10 fold or less.

The chilled water with which the concentrated protein solution is mixedhas a temperature of less than about 15° C., generally about 3° to about15° C., preferably less than about 10° C., since improved yields ofprotein isolate in the form of protein micellar mass are attained withthese colder temperatures at the dilution factors used.

In a batch operation, the batch of concentrated protein solution isadded to a static body of chilled water having the desired volume, asdiscussed above. The dilution of the concentrated protein solution andconsequential decrease in ionic strength causes the formation of acloud-like mass of highly associated protein molecules in the form ofdiscrete protein droplets in micellar form. In the batch procedure, theprotein micelles are allowed to settle in the body of chilled water toform an aggregated, coalesced, dense, amorphous sticky gluten-likeprotein micellar mass (PMM). The settling may be assisted, such as bycentrifugation. Such induced settling decreases the liquid content ofthe protein micellar mass, thereby decreasing the moisture contentgenerally from about 70% by weight to about 95% by weight to a value ofgenerally about 50% by weight to about 80% by weight of the totalmicellar mass. Decreasing the moisture content of the micellar mass inthis way also decreases the occluded salt content of the micellar mass,and hence the salt content of dried isolate.

Alternatively, the dilution operation may be carried out continuously bycontinuously passing the concentrated protein solution to one inlet of aT-shaped pipe, while the diluting water is fed to the other inlet of theT-shaped pipe, permitting mixing in the pipe. The diluting water is fedinto the T-shaped pipe at a rate sufficient to achieve the desireddegree of dilution of the concentrated protein solution.

The mixing of the concentrated protein solution and the diluting waterin the pipe initiates the formation of protein micelles and the mixtureis continuously fed from the outlet from the T-shaped pipe into asettling vessel, from which, when full, supernatant is permitted tooverflow. The mixture preferably is fed into the body of liquid in thesettling vessel in a manner which minimizes turbulence within the bodyof liquid.

In the continuous procedure, the protein micelles are allowed to settlein the settling vessel to form an aggregated, coalesced, dense,amorphous, sticky, gluten-like protein micellar mass (PMM) and theprocedure is continued until a desired quantity of the PMM hasaccumulated in the bottom of the settling vessel, whereupon theaccumulated PMM is removed from the settling vessel. In the batchprocess, the settling may be assisted, such as by centrifugation.

The combination of process parameters of concentrating of the proteinsolution to a preferred protein content of at least about 200 g/L andthe use of a dilution factor less than about 15, result in higheryields, often significantly higher yields, in terms of recovery ofprotein in the form of protein micellar mass from the original mealextract, and much purer isolates in terms of protein content thanachieved using any of the known prior art protein isolate formingprocedures discussed in the aforementioned U.S. patents.

By the utilization of a continuous process for the recovery of canolaprotein isolate as compared to the batch process, the initial proteinextraction step can be significantly reduced in time for the same levelof protein extraction and significantly higher temperatures can beemployed in the extraction step. In addition, in a continuous operation,there is less chance of contamination than in a batch procedure, leadingto higher product quality and the process can be carried out in morecompact equipment.

The settled isolate is separated from the residual aqueous phase orsupernatant, such as by decantation of the residual aqueous phase fromthe settled mass or by centrifugation. The PMM may be used in the wetform or may be dried, by any convenient technique, such as spray drying,freeze drying or vacuum drum drying, to a dry form. The dry PMM has ahigh protein content, in excess of about 90 wt % protein, preferably atleast about 100 wt % protein (calculated as Kjeldahl N×6.25), and issubstantially undenatured (as determined by differential scanningcalorimetry). The dry PMM isolated from fatty oil seed meal also has alow residual fat content, when the procedures of U.S. Pat. Nos.5,844,086 and 6,005,076 are employed as necessary, which may be belowabout 1 wt %. The canola protein isolate contains decreased quantitiesof phytic acid, when compared to extraction of meal with aqueous sodiumchloride solution under the same reaction conditions, and whichpreferably may be below about 1 wt %.

The supernatant from the PMM formation and settling step containssignificant amounts of canola protein, not precipitated in the dilutionstep, and is processed to recover canola protein isolate therefrom. Thesupernatant from the dilution step, following removal of the PMM, isconcentrated to increase the protein concentration thereof. Suchconcentration is effected using any convenient selective membranetechnique, such as ultrafiltration, using membranes with a suitablemolecular weight cut-off permitting low molecular weight species,including the salt and other non-proteinaceous low molecular weightmaterials extracted from the protein source material, to pass throughthe membrane, while retaining canola protein in the solution.Ultrafiltration membranes having a molecular weight cut-off of about3,000 to 100,000 daltons, having regard to differing membrane materialsand configuration, may be used. Concentration of the supernatant in thisway also reduces the volume of liquid required to be dried to recoverthe protein. The supernatant generally is concentrated to a proteinconcentration of about 100 to about 400 g/L, preferably about 200 toabout 300 g/L, prior to drying. Such concentration operation may becarried out in a batch mode or in a continuous operation, as describedabove for the protein solution concentration step.

The concentrated supernatant may be dried by any convenient technique,such as spray drying, freeze drying or vacuum drum drying, to a dry formto provide a further canola protein isolate. Such further canola proteinisolate has a high protein content, in excess of about 90 wt %,preferably at least about 100 wt % protein (calculated as KjeldahlN×6.25) and is substantially undenatured (as determined by differentialscanning calorimetry).

If desired, at least a portion of the wet PMM may be combined with atleast a portion of the concentrated supernatant prior to drying thecombined protein streams by any convenient technique to provide acombined canola protein isolate composition according to one invention.The relative proportions of the proteinaceous materials mixed togethermay be chosen to provide a resulting canola protein isolate compositionhaving a desired profile of 2S/7S/12S proteins. Alternatively, the driedprotein isolates may be combined in any desired proportions to provideany desired specific 2S/7S/12S protein profiles in the mixture andthereby provide a composition according to the invention. The combinedcanola protein isolate composition has a high protein content, in excessof about 90 wt %, preferably at least about 100 wt %, (calculated asKjeldahl N×6.25) and is substantially undenatured (as determined bydifferential scanning calorimetry).

In another alternative procedure, where a portion only of theconcentrated supernatant is mixed with a part only of the PMM and theresulting mixture dried, the remainder of the concentrated supernatantmay be dried as may any of the remainder of the PMM. Further, dried PMMand dried supernatant also may be dry mixed in any desired relativeproportions, as discussed above.

By operating in this manner, a number of canola protein isolates may berecovered, in the form of dried PMM, dried supernatant and driedmixtures of various proportions by weight of PMM-derived canola proteinisolate and supernatant-derived canola protein isolate, generally fromabout 5:95 to about 95:5 by weight, which may be desirable for attainingdiffering functional and nutritional properties based on the differingproportions of 2S/7S/12S proteins in the compositions.

As an alternative to dilution of the concentrated protein solution intochilled water and processing of the resulting precipitate andsupernatant as described above, protein may be recovered from theconcentrated protein solution by dialyzing the concentrated proteinsolution to reduce the salt content thereof. The reduction of the saltcontent of the concentrated protein solution results in the formation ofprotein micelles in the dialysis tubing. Following dialysis, the proteinmicelles may be permitted to settle, collected and dried, as discussedabove. The supernatant from the protein micelle settling step may beprocessed, as discussed above, to recover further protein therefrom.Alternatively, the contents of the dialysis tubing may be directlydried. The latter alternative procedure is useful where small laboratoryscale quantities of protein are desired.

EXAMPLES Example 1

This Example describes the preparation of canola oil seed meal and thesubsequent processing to obtain a canola protein isolate.

125 kg of canola seed of the variety Argentina was processed accordingto the process depicted in FIG. 1. The seed was first submitted to heattreatment in a cooker heated by steam at 90° C. for a 10 minute holdtime in order to deactivate myrosinase and other enzymes. After coolingthe resulting 115.8 kg of inactivated canola oil in a fluid bed dryer,the seeds were cracked and the hulls were partially removed by airaspiration.

The larger canola meats (overs) were separated with a 14-mesh vibratoryscreen and the overs were recycled 4 times to the cracking mill toprovide 42.4 kg of mainly canola meats and a smaller fraction of hulls.The unders (36 kg) were passed through final air aspiration for removalof residual hulls. The final meats (35.3 kg) or the unders fraction,were flaked by a flaking mill before passage of 34.1 kg of canola flakesto a Soxhlet extractor for oil extraction by solvent while the oversfraction was discarded.

The dehulled and defatted meal from the oil extraction (16.17 kg) wasused as the starting material for protein extraction, as described inExample 2 below. The dehulled canola meal was identified as SD024.

Two additional fractions of dehulled and defatted canola meal wereobtained from a second 130.4 kg batch of canola seed of the varietyArgentina following the procedure of FIG. 2. For this batch, the seedswere initially cracked and the hulls partially removed by airaspiration.

The larger canola meats were separated with a 14-mesh vibratory screenand the overs were recycled 4 times to provide 52.2 kg of canola meatsand hulls. After the last pass through the vibratory screen, both unders(49.2 kg) and overs were heat-treated using steam at 90° C. for 10minutes. The fractions were cooled down in a fluidized bed dryer. Thefinal meats were flaked in a flaking mill. The flakes obtained from theunders (38.1 kg) were solvent extracted directly using a Soxhletextractor to remove the oil, producing (11.35 kg) a defatted mealidentified as SD029. The flakes obtained from the overs were airaspirated another time and the aspirated flakes solvent extracted usinga Soxhlet extractor to remove the oil, producing a defatted meal (11.37kg) identified as SD027.

The temperature profiles during inactivation of the canola oil seed forsamples SD024 (“Batch #1”), SD029 (“Batch #2 unders”) and SD027 (“Batch#2 overs”) are shown in FIG. 4.

In the procedure, a total of 35.3 kg of dehulled meats (unders) wasrecovered from 112.3 kg of inactivated canola seed in batch #1 toproduce a total yield of 31.43 wt %. A total of 38.1 kg of dehulled andflaked fines (unders) were produced from 130.4 kg of batch #2 canola,resulting in a yield of 29.2 wt %. The relatively low yields of dehulledcanola can partly be attributed to ineffective cracking of the smallercanola seeds due to the use of coarse rollers in the cracking mill. Theuse of finer pitch rolls (18 corrugations per inch) will permit anarrower gap between the rolls and enable cracking of smaller seeds. Alarger and more uniform seed may also increase the yield and consistencyof dehulling.

Aspiration conditions were adjusted in order to achieve effectiveseparation of hulls from the meats. The differential air pressuresetting of 0.4 to 0.8 inches of water resulted in an effectiveseparation. Larger pressure differentials caused excessive endosperm tobe removed with the hull fraction.

The meat fraction recovered from air aspiration consisted of a widerange of particle size and the canola that was more finely crackedcontained a smaller proportion of hull fragments. As a result, thesmaller dehulled meats fraction could be recovered from the larger meatsand hulls by screening through the 14 mesh vibratory screen. The optimumscreen size was pre-selected by hand screening tests prior to set-up ofthe equipment.

Flaking was carried out to rupture the oil cells by passing the dehulledendosperm fractions through a set of smooth rollers on a Lauhauf flakingmill.

The dehulled meat from both batches #1 and #2 were effectively flakedusing a gap setting of 0.08 mm and produced a flake thickness rangingfrom 0.101 to 0.125 mm. Flakes produced from the batch #2 process,however, were fragile and crumbled somewhat in comparison to the batch#1 flakes. This result indicated that inactivating the canola seed priorto dehulling produced a more stable flake.

Following defatting, residual oil content of the batch #1 defattedcanola meal was 1.50 wt %. Batch #2 meal contained 1.87 wt % and 1.23 wt% oil in the unders and overs fraction, respectively.

Example 2

This Example illustrates the preparation of canola protein isolates fromthe defatted meals prepared according to the procedures of Example 1.

Dehulled, defatted and myrosinase-inactivated canola meals, prepared asdescribed in Example 1, were processed according to the procedure ofFIG. 3, to produce canola protein isolates.

‘a’ kg of dehulled, defatted and myrosinase inactivated canola meal wasadded to ‘b’ L of 0.15 M NaCl solution at ambient temperature andagitated for 30 minutes to provide an aqueous protein solution. Theresidual canola meal was removed by filtration through cheese cloth orby other suitable filtration methods. The resulting protein solution wasclarified by centrifugation to produce ‘c’ L of a clarified proteinsolution having a protein content of ‘d’ g/L.

A ‘e’ L aliquot of the protein extract solution was reduced in volume to‘f’ L by concentration on an ultrafiltration system using ‘g’ daltonmolecular weight cutoff membrane. The resulting concentrated proteinsolution had a protein content of ‘h’ g/L. The concentrated proteinsolution was then diafiltered using ‘i’ dalton molecular cut-offmembranes using ‘j’ L of 0.15 M sodium chloride solution containing 0.05wt % ascorbic acid to a final volume of ‘k’ L of diafiltered proteinsolution with a protein content of ‘1’ g/L.

The diafiltered protein solution at ‘m’° C. was diluted ‘n’ into ‘o’° C.water. A white cloud formed immediately and was allowed to settle. Theupper diluting water was removed and the precipitated, viscous, stickymass (PMM) was recovered from the bottom of the vessel in a yield of ‘p’wt % of the extracted protein. The dried PMM derived protein was foundto have a protein content of ‘q’ % (N×6.25) d.b. The product was givendesignation ‘r’.

The parameters ‘a’ to ‘r’ are given in the following Table I:

TABLE I BW-SD024-B03-03A BW-SD029-B10-03A BW-SD027-B17- r C300 C300 02AC300 a 5 5 5 b 50 50 50 c 38.3 39 36 d 25.7 21.6 23.1 e 38.3 39 36 f 2.53.5 2.5 g 10000 10000 10000 h 218.3 218.9 232.0 i 10000 10000 10000 j 5035 17.5 k 1.8 3.5 2.5 l 266.7 218.9 232.0 m 30.5 31 31.4 n 1:10 1:101:10 o 1.7 2 2.2 p 40.2 55.6 57.3 q 106.7 110.1 107.6

The removed diluting water was reduced in volume by ultrafiltrationusing a ‘s’ dalton molecular weight cut-off membrane to a proteinconcentration of ‘t’ g/L. The concentrate was dried. With the additionalprotein recovered from the supernatant, the overall protein recovery was‘u’ wt % of the extracted protein. The dried protein formed had aprotein content of ‘v’ wt % (N×6.25) d.b.

The product was given designation ‘w’. The parameters s to w are givenin the following Table II:

TABLE II BW-SD024-B03-03A BW-SD029-B10-03A BW-SD027-B17- w C200 C200 02AC200 s 10000 10000 10000 t 20.7 52.1 118.0 u 46.7 70.6 78.6 v 103.8103.6 106.2

Example 3

This Example describes the results obtained by following the proceduresof Example 2.

(a) Extraction and Separation Steps:

Table III below represents the apparent extractabilities for the threedifferent meals. The apparent extractability represents the percentageof protein that could be recovered if the total saline volume could berecovered. However, the recovery can vary due to differences in the mealand/or to different liquid hold-up in the meal. When the actual volumepost clarification processes is taken into account for calculations,then the result is protein yield. The apparent extractability is higherthan 40% for all three cases. For SD024 and SD027 meal, they are in thesame order of magnitude with 47.5 wt % and 46.1 wt %, respectively. Thenumber for the SD029 meal is slightly smaller. The apparentextractability is not significantly influenced by the dehulling and heattreatment process of the meal, as the extractability numbers are in thesame range as for low temperature desolventized or marc meal (data notshown).

TABLE III Apparent extractabilities and protein yields in the postfiltration liquids Apparent extractability Protein yield post (wt %)filtration step (wt %) BW-SD024-B03-03A 47.5% 36.4% BW-SD029-B10-03A41.3% 38.0% BW-SD027-B17-03A 46.1% 33.1%

(b) Ultrafiltration #1 and #2:

The protein recovery (Table IV) for SD029 and SD027 meal is similar tothe values usually observed for other meals for ultrafiltration #1 whenusing PVDF 5 spiral membranes. The lower value of 55 wt % for SD024 mealis due to some protein losses in the permeates. A chromatogram of thepermeate showed a significant amount of 2S protein for the batchBW-SD024-B03-03A. This loss of protein is thought be due to the newnessof the membrane employed.

TABLE IV Protein recoveries and protein yields in retentate forultrafiltration #1 Protein recovery Protein in retentate yield postultrafiltration (wt %) (wt %) BW-SD024-B03-03A 55% 17.78%BW-SD029-B10-03A 72% 27.38% BW-SD027-B17-03A 70% 23.15%

For the Ultrafiltration #2, the protein recovery was 75 wt % (SD024), 90wt % (SD029) and 100 wt % (SD027).

(c) Protein Distribution in Final Products:

Tables V and VI below represent the protein distribution for thefinished PMM-derived isolates and supernatant-derived isolates. Theprotein peaks from the SEC chromatograms were considered as one groupbeing 100 wt %. That means, for example, if there is 80 wt % 7S, then 80wt % of the total peak area of all the protein peaks belongs to 7Sprotein.

TABLE V Protein distribution for PMM derived protein isolates obtainedfrom different meals 12S (wt) 7S (wt) 2S (wt) BW-SD024-B03-03A 17.5%81.3% 1.5% BW-SD029-B10-03A 9.6% 81.3% 9.1% BW-SD027-B17-03A 7.9% 82.4%9.7%

As may be seen the protein distribution in the PMM follows the samepattern that has been observed previously (see copending U.S. patentapplication Ser. No. 10/413,371 filed Apr. 15, 2003 (WO 03/088760),assigned to the assignee hereof and the disclosure of which isincorporated herein by reference) that 7S is the major protein in PMM. Areduced 2S amount and therefore a higher 12S concentration was found forthe PMM obtained from SD024 meal which is due to the protein lossthrough the membrane.

TABLE VI Protein distribution of Supernatant-derived protein isolatesobtained from different meals 12S (wt) 7S (wt) 2S (wt) BW-SD024-B03-03A6.8% 81.7% 11.5% BW-SD029-B10-03A 1.5% 16.7% 82.9% BW-SD027-B17-03A 0.7%9.6% 89.7%

As a result of the 2S loss for the SD024 meal, the product yield as wt %of extracted protein was significantly less than for SD027 or SD029meal. The composition of the supernatant-derived isolate resembles thatof the PMM-derived isolate. For the dilution, there is an insufficientamount of 2S protein remaining in solution in the supernatant and,therefore, 2S is not the major protein component. As 7S is also found insupernatant, but at a lower concentration, the absence of 2S has led to7S being the major protein in supernatant-derived isolate. However, forthe later runs with SD029 and SD027 meal, the composition of thesupernatant-derived isolate composition is found to be within the normalrange that has been previously observed for supernatant-derivedisolates.

The above results indicate that, generally, the dehulling and heattreatment process of the meals does not affect the protein compositionof the canola protein isolates obtained.

(d) Canola Protein Isolate Colour:

Table VII and Table VIII below represent the “L”, “a”, “b” colour valuesfor either the dry product or for reconstituted product, in which drypowder was re-suspended in 0.1 M saline and stirred for about an hour,as measured using a Minolta CR-310 colourimeter for the dry product or aHunter Lab DP-9000 colourimeter for reconstituted. The “L” value, with arange from 0 to 100, represents the lightness of the product (L=100being white). The “a” value (from −60 to +60) represents the green-redcolour space. The more negative the “a” value the greener the product,the more the “a” value tends towards +60 the more red the product. The“b” value (from −60 to +60) represents the blue-yellow colour space. Themore negative the “b” value the bluer the product, the more the “b”value tends towards +60, the more yellow the product.

Comparing the lightness of the dry as well as the reconstitutedproducts, the products obtained from the meal batch which has been heattreated in the seed has the highest L values. These products aresignificantly lighter than the ones obtained from meal batch #2 forwhich the heat treatment occurred only after the cracking of the seeds.This result indicates that myrosinase was active and had enough time tocatalyze the degradation of glucosinolates before it was finallyinactivated. The degradation products of the glucosinolates areconsidered to contribute to the darker colour of the PMM-derived isolateand supernatant-derived isolate from this meal.

The protein isolates obtained from the SD024 meal tends more towardsgreen whereas the “a” value for isolates from SD027 and SD029 havehigher numbers and have a more reddish colour. The dry powders andliquid samples do not show the same trend for the blue-yellow colourspace. For example, the “b” value for the dry product for SD024PMM-derived isolate is the smallest of the three different runs, whereasthe SD024 PMM-derived isolate results in the highest “b” value for theliquid sample. The most yellow powder was observed for the SD027 meal inboth the PMM-derived isolate and the supernatant-derived isolate. Theleast yellow product is obtained from SD024 meal for PMM-derived isolateand for SD029 meal for supernatant-derived isolate.

When looking at the liquid colour analysis, the most yellow ofPMM-derived isolates is the one resulting from SD024 meal, forsupernatant-derived isolate, the most yellow is obtained form SD027.

TABLE VII L, a, b colour values in the dry powdered products PMM IsolateSupernatant Isolate L a b L a b BW-SD024-B03-03A 85.36 −1.57 +21.3487.06 −1.40 +18.24 BW-SD029-B10-03A 74.76 +0.15 +24.69 83.02 −0.61+15.94 BW-SD027-B17-03A 79.07 +0.25 +27.26 83.58 −0.44 +21.18

TABLE VIII L, a, b colour values in the liquid of reconstituted productsPMM Isolate Supernatant Isolate L a b L a b BW-SD024-B03-03A 51.18 −0.47+21.49 47.30 +0.32 +16.08 BW-SD029-B10-03A 30.67 +0.34 +13.22 21.84+7.90 +13.47 BW-SD027-B17-03A 27.92 +5.38 +14.72 25.99 +11.20 +16.75

Example 4

This Example describes the enzyme inactivation employing radio frequencytreatment.

A batch of canola seed having a moisture content of about 9% seed wassplit into three 2 kg samples. One of the samples served as control andwas not treated further.

Two 2 kg samples of canola seed of were exposed to radio frequencytreatment. The exposure to radio frequency results in a rapid increaseof temperature over the whole volume of the canola seed sample. Onesample was heated within about 160 seconds from ambient temperature to90° C. and hold at 90° C. for 5 minutes. The second sample was heatedfrom ambient temperature to 90° C. within about 160 seconds and was thenhold at 90° C. for 10 minutes.

After the holding at 90° C., both samples were cooled down to 30° C. bybeing spread out on a baking pan and stored in a cooling room at 4° C.for about 10 minutes.

Myrosinase activity was tested by testing for glucose, a breakdownproduct of the glucosinolate degradation. The test procedure is asfollows: an 100 g aliquot of canola seed is homogenized in 250 ml tapwater with a Silverson homogenizer at 6000 rpm until the mixture forms apaste. This mixture is allowed to sit for 20 minutes and is thencentrifuged at 10000×g for 5 minutes. The supernatant from this step isdecanted and tested for glucose employing Diastix glucose monitoringstrips (Bayer).

All three seed samples, heat treated and control, were tested forglucose. The results are presented in Table IX below.

TABLE IX Glucose Level in Supernatant Control   6 mmol/l Canola seedheated for 5 minutes at 90° C. <5 mmol/l Canola seed heated for 10minutes at 90° C. Not detected

There is no glucose detected for the canola seed sample that was heattreated at 90° C. for 10 minutes. This shows that employing radiofrequency is an effective means to inactivate myrosinase enzyme.

Example 5

This Example illustrates the preparation of enzyme inactivated canolameal to be used for the production of protein isolates samples insufficient amount to carry out sensory analysis.

Three tons of canola seed were continuously processed to prepare anenzyme inactivated canola meal. The deactivation of the enzyme was doneusing a two tray Simon-Rosedown cooker. The cooker was preheated priorto the start of the run. Steam pressures were adjusted while running tomaintain the desired seed temperatures. Temperatures in the trays were60° C. (±5° C.) for the top tray and 82 to 86° C. for the bottom tray.The feed rate of canola seed to the cooker was ˜300 kg/hr and theresidence time in the bottom tray was ˜12 minutes. The deactivated seedwas then transferred to a grain dryer and quickly cooled to <60° C.

After deactivation, the canola seed was too dry and required tempering.The seed moisture was 5.74% and was tempered by spraying the seed with3% water (w/w) to raise the moisture content to ˜8.0%. The water andseed were blended for approximately 15 minutes and then transferred intoa portabin, covered and allowed to equilibrate for a minimum of 12hours.

Flaking was done to rupture oil cells and prepare a thin flake with alarge surface area for cooking/pre-pressing by passing the seed througha flaking mill. The flake thickness was between 0.18-0.23 mm. The feedrate was controlled to balance the rate of pressing and wasapproximately 130 kg/hr.

Cooking was done to further rupture oil cells, make flakes pliable andincrease the efficiency of the expeller by lowering the viscosity of theoil contained. The cooker was preheated prior to the start of the run.Steam pressures were adjusted while running to maintain the desiredflake temperatures. Temperatures in the trays were 42° C. (±2° C.) forthe top tray and 65° C. (±3° C.) for the bottom tray.

Pressing removed approximately ⅔ to ¾ of the oil and prepared a materialsuitable for solvent extraction. The material requires crush resistanceto hold up in the extractor and porosity for good mass transfer anddrainage. The flaked and cooked seed was pressed using a Simon-Rosedownpre-press. The crude press oil was discarded.

Solvent extraction was the contacting of press cake with iso-hexane toremove the oil from the cake mass. Two mechanisms were in operation: theleaching of the oil into the solvent, and the washing of the marc meal(iso-hexane-solids) with progressively weaker miscelles (hexane-oil).Extraction is normally a continuous counter-current process.

The canola seed press cake was extracted on a Crown Iron Works LoopExtractor (Type II) with iso-hexane using a total residence time ofapproximately 100 minutes (loop in to loop out) and a solvent to solidsratio of approximately 3.2:1 (w:w). The crude oil was desolventized in arising film evaporator and steam stripper. The oil was discarded.

Desolventization of the marc (hexane-solids) was done in a steamjacketed Schnecken screw and 2 tray desolventizer-toaster. Thetemperatures in the trays were <50° C. at Schnecken Exit, 50° C. (±5°C.) for the desolventizer tray and 45° C. (±5° C.) for the toastingtray.

Vacuum drying was done to finish the desolventization of the extractedcanola meal. Approximately 150 kg per batch of defatted canola meal wasloaded into a Littleford Reactor. The meal was then heated to 47° C.(±2° C.) under a vacuum of 23-25 mmHG. The meal was held at thistemperature for 2 hours, then discharged into plastic lined fiber drums.A total of 1317.3 kg of enzyme inactivated, defatted and vacuumdesolventized canola meal was produced.

Example 6

This Example illustrates the preparation of canola protein isolates fromthe defatted, enzyme inactivated meal according to Example 5 and fromcommercially-available low temperature desolventized meal. The canolaprotein isolates will be used to compare colour and flavour.

The defatted, enzyme inactivated meal according to Example 5 was giventhe designation SA034 and the commercial meal was given the designationAL022.

‘a’ kg of canola meal was added to ‘b’ L of 0.15 M NaCl solution atambient temperature, agitated for 30 minutes to provide an aqueousprotein solution. The residual canola meal was removed by vacuumfiltration (in the case of BW-AL022-B24-03A) or decanter centrifugation(in the case of BW-SA034-E06-04A C300) and disc centrifugation. Theresulting protein solution was clarified by filter press filtration toproduce ‘c’ L of a clarified protein solution having a protein contentof ‘d’ g/L.

A ‘e’ L aliquot of the protein extract solution was reduced in volume to‘f’ L by concentration on an ultrafiltration system using ‘g’ daltonmolecular weight cutoff membranes. The resulting concentrated proteinsolution had a protein content of ‘h’ g/L. The concentrated proteinsolution then was diafiltered on a diafiltration system using ‘i’ Daltonmolecular weight cut-off membranes using ‘j’ L of ‘k’ M NaCl solutioncontaining 0.05 wt % ascorbic acid to a final volume of ‘l’ L with aprotein content of ‘m’ g/L.

The concentrated solution at ‘n’° C. was diluted ‘o’ into ‘p’° C. water.A white cloud formed immediately and was allowed to settle. The upperdiluting water was removed and the precipitated, viscous, sticky mass(PMM) was recovered from the bottom of the vessel in a yield of ‘q’ wt %of the extracted protein. The dried PMM derived protein was found tohave a protein content of ‘r’ % (N×6.25) d.b. The product was givendesignation ‘s’.

BW-AL022-B24-03A C300 BW-SA034-E06-04A C300 a 150 150 b 1000 1500 c 11801265 d 12.2 15.7 e 1180 1265 f 45 65 g 10000 5000 h 283 213 i 10000 5000j 235 325 k 0.15 0.1 l 35.35 57.5 m 316 248 n 31.9 29.6 o 1:15 1:10 p3.7 3.1 q 48.7 33.6 r 102.8 100.9

The removed diluting water was reduced in volume by ultrafiltrationusing a ‘t’ dalton molecular weight cut-off membrane to a proteinconcentration of ‘u’ g/L. The concentrate was dried. With the additionalprotein recovered from the supernatant, the overall protein recovery was‘v’ wt % of the extracted protein. The dried protein formed had aprotein content of ‘w’ wt % (N×6.25) d.b.

The product was given designation ‘x’.

x BW-AL022-B24-03A C200 BW-SA034-E06-04A C200 t 10000 100000 u 158.7192.1 v 78.2 56.4 w 104.4 94.7

Example 7

This Example describes the results obtained by following the procedureof Example 6.

(a) Sensory Analysis

The canola protein isolate samples were submitted for sensory analysis.The sensory panel consisted of 11 trained panelists. Each panelist wasasked which sample has the least amount of flavour and which sample thepanelist would prefer.

The canola protein isolates obtained by following the procedure ofExample 6 were resuspended in 0.05M saline solution at a concentrationof 5% w/v. The protein powders were completely solubilized beforesensory tests started.

Table X below shows the results of the sensory analysis for PMMproducts. It appears that the isolate derived from the enzymeinactivated meal, were found to be the ones with the least amount offlavour and were also the more preferred products. 64% of the panelistfound that the PMM of the enzyme inactivated meal had the least amountof flavour whereas 27% found the low temperature meal derived PMM tohave the least amount of flavour. 9% of the panelist could not find adifference in between the two products.

When being asked which product they would prefer, 64% of the panelistgave the PMM derived from the enzyme inactivated meal as theirpreference, 18% preferred the low temperature meal derived product and18% did not prefer either one of the products.

TABLE X Sensory analysis of C300 products Had the least amount offlavour Preferred Product BW-AL022-B24-03A C300 3 2 BW-SA034-E06-04AC300 7 7 Could not find a difference 1 2

Table XI below shows the results of the sensory analysis for supernatantderived protein isolates. It appears that the isolate derived from theenzyme inactivated meal, were found to be the ones with the least amountof flavour and were also the more preferred products. 55% of thepanelist found that the supernatant derived protein of the enzymeinactivated meal had the least amount of flavour whereas 27% found theproduct obtained from low temperature meal to have the least amount offlavour. 9% of the panelist could not find a difference in between thetwo products.

When being asked which product they would prefer, 82% of the panelistgave the supernatant derived protein obtained from the enzymeinactivated meal as their preference, 9% preferred the low temperaturemeal derived product and 9% did not prefer either one of the products.

TABLE XI Sensory analysis of C200 products Had the least amount offlavour Preferred Product BW-AL022-B24-03A C200 3 1 BW-SA034-E06-04AC200 6 9 Could not find a difference 2 1

(a) Colour Analysis

Table XII below shows the “L”, “a”, “b” colour values for reconstitutedproduct (5% w/v product in 0.05M saline) as measured using a Hunter LabD9000 colourimeter. The “L value, with a range from 0 to 100, representsthe lightness of the product (L=100 being white). The “a” value (from−60 to +60) represents the green-red colour space. The more negative the“a” value the greener the product, the more the “a” value tends towards+60 the more red the product. The “b” value (from −60 to +60) representsthe blue-yellow colour space. The more negative the “b” value the bluerthe product, the more the “b” value tend towards +60 the more yellow theproduct.

Comparing the lightness of the liquid samples it appears that for bothprotein isolates, PMM and supernatant derived, the L value wassignificantly higher for the enzyme inactivated meal derived productsthan for the low temperature meal derived products. That means that theenzyme inactivated meal produced in both cases a much lighter proteinisolate.

For the red-green colour space as well as the blue-yellow colour spaceboth the PMM and the supernatant isolate follow the same trend. Usingthe enzyme inactivated meal as starting material the “a” value isslightly decreased compared to low temperature meal, meaning that thesamples tend more towards a greenish colour. The “b” value increaseswhen using enzyme inactivated meal, meaning that the samples appear moreyellow compared to samples obtained from low temperature meal.

TABLE XII L, a, b colour values in the liquid of reconstituted productsPMM Isolate Supernatant Isolate L a b L a b BW-SA034-E06-04A 47.11 3.2526.69 39.62 2.74 20.58 BW-AL022-B24-03A 32.09 6.79 18.94 23.0 7.8 12.41

SUMMARY OF DISCLOSURE

In summary of this disclosure, the present invention provided a processof producing a canola protein isolate of improved colour and taste byinitially heat-inactivating myrosinase and other enzymes in the canolaoil seeds prior to further processing of the oil seeds. Modificationsare possible within the scope of this invention.

1. A method of forming a canola protein isolate having a protein contentof at least about 90 wt % from intact canola seeds which comprises: heattreating the intact canola oil seeds to deactivate enzymes therein,dehulling the canola oil seeds, removing canola oil from the heattreated and dehulled oil seeds to provide a canola oil seed meal, andprocessing the canola oil seed meal to recover therefrom the canolaprotein isolate, wherein said canola oil seed meal is processed torecover said canola protein isolate by the steps of: (i) extracting thecanola oil seed meal with an aqueous salt solution to causesolubilization of canola protein in said the canola protein seed meal toform an aqueous canola protein solution having a pH of about 5 to about6.8, (ii) separating the aqueous protein solution from residual canolaoil seed meal, (iii) increasing the protein concentration of saidaqueous protein solution while maintaining the ionic strengthsubstantially constant by using a selective membrane technique toprovide a concentrated protein solution, (iv) diluting the concentratedcanola protein isolate into chilled water having a temperature of belowabout 15° C. to cause the formation of discrete protein particles in theaqueous phase in the form of micelles, (v) settling the protein micellesto form an amorphous, sticky, gelatinous, gluten-like protein micellarmass, (vi) recovering the protein micellar mass from supernatant, and(vii) processing the supernatant, on a batch, semi-continuous orcontinuous basis, to recover additional quantities of canola proteinisolate therefrom.
 2. The method of claim 1 wherein said heat treatedand dehulled oil seeds are flaked prior to said oil removal step.
 3. Themethod of claim 1, wherein said canola oil seed meal is produced by:heat treating the intact canola oil seeds to inactivate enzymes therein,cooling the heat treated canola oil seeds, cracking the hulls of theheat treated canola oil seeds, removing cracked hulls from canola seedsto produce canola meats, and removing canola oil from the canola meatsby solvent extraction to leave a meal.
 4. The method of claim 3 whereinan overs fraction and an unders fraction are separated from the crackedhulls, the overs fraction is recycled to the cracking and separationsteps, the unders fraction is subjected to air aspiration for furtherremoval of hulls and the recycled overs fraction and/or air aspiratedunders fraction are flaked prior to said solvent extraction step.
 5. Themethod of claim 1 wherein the canola oil seed meal is processed torecover therefrom a canola protein isolate having a protein content ofat least about 100 wt % (N×6.25).
 6. The method of claim 1 wherein saidinactivation is effected by heating using steam.
 7. The method of claim1 wherein said inactivation is effected by heating using radio frequencyradiation.
 8. The method of claim 1, wherein said steps (i) to (vi) areeffected on a batch basis and said extracting of said oil seed meal iseffected using an aqueous salt solution having an ionic strength of atleast about 0.10 and a pH of about 5 to about 6.8 and said aqueousprotein solution has a protein content of about 5 to about 40 g/L. 9.The method of claim 1, wherein said steps (i) to (vi) are effected on acontinuous basis and said extraction step is effected by: (i)continuously mixing an oil seed meal with an aqueous salt solutionhaving an ionic strength of at least about 0.10 and a pH of about 5 toabout 6.8 at a temperature of about 5° to about 65° C., and (ii)continuously conveying said mixture through a pipe while extractingprotein from the oil seed meal to form an aqueous protein solutionhaving a protein content of about 5 to about 40 g/L in a period of timeup to about 10 minutes.
 10. The method of claim 1, wherein followingsaid separating of the aqueous protein solution from the residual canolaseed meal, the aqueous protein solution is subjected to a pigmentremoval step.
 11. The method of claim 10, wherein said pigment removalstep is effected by daifiltration of the aqueous protein solution. 12.The method of claim 10, wherein said pigment removal step is effected bymixing a pigment adsorbing agent with the aqueous protein solution andsubsequently removing the pigment adsorbing agent from the aqueousprotein solution.
 13. The method of claim 1, wherein said oil seed mealis extracted with water and subsequent thereto salt is added to theresulting aqueous protein solution to provide an aqueous proteinsolution having an ionic strength of at least about 0.10.
 14. The methodof claim 1, wherein said concentration step is effected byultrafiltration to produce a concentrated protein solution having aprotein content of at least about 200 g/L.
 15. The method claim 1,wherein said concentrated protein solution is subjected to diafiltrationusing an aqueous salt solution of the same molarity and pH as theextracting solution.
 16. The method of claim 15, wherein saiddiafiltration is effected until no further quantities of phenolics andvisible colour are present in the permeate.
 17. The method of claim 15,wherein said anti-oxidant is present in the diafiltration medium duringat least part of the diafiltration step.
 18. The method of claim 1,wherein the concentrated protein solution, optionally diafiltered, issubjected to a pigment removal step.
 19. The method of claim 1, whereinthe concentrated protein solution, optionally diafiltered, is subjectedto a pasteurization step.
 20. The method of claim 1, wherein therecovered protein micellar mass is dried to a proteinaceous powder. 21.The process of claim 1 wherein said additional quantities of proteinisolate are recovered from the supernatant by concentrating thesupernatant to a protein concentration of about 100 to about 400 g/L anddrying the concentrated supernatant.
 22. The process of claim 21 whereinsaid supernatant is concentrated to a concentration of about 200 toabout 300 g/L.
 23. The process of claim 1 wherein said additionalquantities of protein isolate are recovered from the supernatant byconcentrating the supernatant to a protein concentration of about 100 toabout 400 g/L mixing the concentrated supernatant with the recoveredprotein micellar mass, and drying the mixture.
 24. The process of claim23 wherein said supernatant is concentrated to a concentration of about200 to about 300 g/L.
 25. The process of claim 1 wherein said additionalquantities of protein isolate are recovered from the supernatant byconcentrating the supernatant to a protein concentration of about 100 toabout 400 g/L mixing a portion of said concentrated supernatant with atleast a portion of the recovered protein micellar mass, and drying theresulting mixture.
 26. The process of claim 25 wherein the remainder ofthe concentrated supernatant is dried and any remainder of the recoveredprotein micellar mass is dried.
 27. The process of claim 25 wherein saidsupernatant is concentrated to a concentration of about 200 to about 300g/L.